Mixotrophic growth of Thiobacillus A2 on acetate and thiosulfate as growth limiting substrates in the chemostat

During heterotrophic growth on acetate, in batch culture, the autotrophic growth potential of Thiobacillus A2, i.e. the capacity to oxidize thiosulfate and to fix carbon dioxide via the Calvin cycle, was completely repressed. The presence of thiosulfate in a batch culture with acetate as the organic substrate partly released the repression of the thiosulfate oxidizing system. Cultivation of the organism in continuous culture at a dilution rate of 0.05 h^-1 with different concentration ratios of thiosulfate and acetate in the reservoir medium led to mixotrophic growth under dual substrate limitation. Growth on the different mixtures of acetate and thiosulfate yielded upto 30% more cell dry weight than predicted from the growth yields on comparable amounts of these substrates separately. The extent to which the carbon dioxide fixation capacity and the maximum thiosulfate and acetate oxidation capacity are repressed appeared to be a function of the thiosulfate to acetate concentration ratio in the reservoir medium. The results of ¹⁴C-acetate assimilation experiments and of gas-analysis demonstrated that the extent to which acetate was assimilated depended also on the substrate ratio in the inflowing medium. Under the different growth conditions surprisingly little variation was found in some tri-carboxylic acid cycle enzyme activities. Cultivation of T. A2 at different growth rates with a fixed mixture of thiosulfate (18 mM) and acetate (11 mM) in the medium, showed that dual substrate limitation occured at dilution rates ranging from 0.03–0.20 h^-1.Abbreviations PPO 2,5-diphenoloxazol - RubPCase Ribulose-1,5-bisphophate carboxylase - Tris tris (hydroxymethyl) aminomethane - EDTA ethylenediaminetetra-acetic acid     

Microbial sulfide oxidation in the oxic–anoxic transition zone of freshwater sediment: involvement of lithoautotrophic Magnetospirillum strain J10

The oxic–anoxic transition zone (OATZ) of freshwater sediments, where opposing gradients exist of reduced iron and sulfide with oxygen, creates a suitable environment for microorganisms that derive energy from the oxidation of iron or sulfide. Gradient microcosms incubated with freshwater sediment showed rapid microbial turnover of sulfide and oxygen compared with sterile systems. Microcosms with FeS as a substrate also showed growth at the OATZ and subsequent dilution series resulted in the isolation of three novel strains, of which strain J10 grows chemolithoautotrophically with reduced sulfur compounds under microaerobic conditions. All three strains are motile spirilla with bipolar flagella, related to the genera Magnetospirillum and Dechlorospirillum within the Alphaproteobacteria . Strain J10 is closely related to Magnetospirillum gryphiswaldense and is the first strain in this genus found to be capable of autotrophic growth. Thiosulfate was oxidized completely to sulfate, with a yield of 4 g protein mol⁻¹ thiosulfate, and autotrophic growth was evidenced by incorporation of ¹³C derived from bicarbonate into biomass. A putative gene encoding ribulose 1,5-bisphosphate carboxylase/oxygenase type II was identified in strain J10, suggesting that the Calvin–Benson–Bassham cycle is used for autotrophic growth. Analogous genes are also present in other magnetospirilla, and in the autotrophically growing alphaproteobacterium magnetic vibrio MV-1.     

Chemolithoautotrophy and mixotrophy in the thiophene-2-carboxylic acid-utilizing Xanthobacter tagetidis

Xanthobacter tagetidis grew as a chemolithotrophic autotroph on thiosulfate and other inorganic sulfur compounds, as a heterotroph on thiophene-2-carboxylic acid, acetic acid and α-ketoglutaric acid, and as a mixotroph on thiosulfate in combination with thiophene-2-carboxylic acid and/or acetic acid. Autotrophic growth on one-carbon organosulfur compounds, and intermediates in their oxidation are also reported. Thiosulfate enhanced the growth yields in mixotrophic cultures, presumably by acting as a supplementary energy source, since ribulose bisphosphate carboxylase was only active in thiosulfate-grown cells and was not detected in mixotrophic cultures using thiosulfate with thiophene-2-carboxylic acid. Bacteria grown on thiophene-2-carboxylic acid also oxidized sulfide, thiosulfate and tetrathionate, indicating these as possible sulfur intermediates in thiophene-2-carboxylic acid degradation. Thiosulfate and tetrathionate were oxidized completely to sulfate and, consequently, did not accumulate as products of thiophene-2-carboxylic acid oxidation in growing cultures. K _m and V _max values for the oxidation of thiosulfate, tetrathionate or sulfide were 13 μM and 83 nmol O₂ min^–1 (mg dry wt.)^–1, respectively; thiosulfate and tetrathionate became autoinhibitory at concentrations above 100 μM. The true growth yield (Y_max) on thiophene-2-carboxylic acid was estimated from chemostat cultures (at dilution rates of 0.034–0.094 h^–1) to be 112.2 g mol^–1, with a maintenance coefficient (m) of 0.3 mmol thiophene-2-carboxylic acid (g dry wt.)^–1 h^–1, and the maximum specific growth rate (μ_max) was 0.116 h^–1. Growth in chemostat culture at a dilution rate of 0.041 h^–1 indicated growth yields [g dry wt. (mol substrate)^–1] of 8.1 g (mol thiosulfate)^–1, 60.9 g (mol thiophene-2-carboxylic acid)^–1, and 17.5 g (mol acetic acid)^–1, with additive yields for growth on mixtures of these substrates. At a dilution rate of 0.034 h^–1, yields of 57.8 g (mol α-ketoglutaric acid)^–1 and 60.7 g (mol thiophene-2-carboxylic acid)^–1 indicated some additional energy conservation from oxidation of the thiophene-sulfur. SDS-PAGE of cell-free preparations indicated a polypeptide (M _r, 21.0 kDa) specific to growth on thiophene-2-carboxylic acid for which no function can yet be ascribed: no metabolism of thiophene-2-carboxylic acid by cell-free extracts was detected. It was shown that X. tagetidis exhibits a remarkable degree of metabolic versatility and is representative of facultatively methylotrophic and chemolithotrophic autotrophs that contribute significantly to the turnover of simple inorganic and organic sulfur compounds (including substituted thiophenes) in the natural environment. Received: 1 July 1997 / Accepted: 3 November 1997     

Capacity of chromatiaceae for chemotrophic growth. Specific respiration rates of Thiocystis violacea and Chromatium vinosum

The capacity for chemoautotrophic, mixotrophic and organotrophic growth in the dark was tested with 45 strains of 17 species (11 genera) of the Chromatiaceae. The auxanographic deep agar shake culture method was used; the gas phase contained 5% O₂ and 1% CO₂ in N₂. All strains tested of Chromatium vinosum, C. minus, C. violascens, C. gracile, Thiocystis violacea, Amoebobacter roseus, Thiocapsa roseopersicina gave positive growth responses under chemoautotrophic and mixotrophic conditions (extra carbon source acetate); one strain of Thiocapsa roseopersicina grew also organotrophically on acetate alone. No growth was obtained with the remaining 17 strains of ten species. None of the five type species (three genera) of the Chlorobiaceae grew under chemotrophic conditions. With Thiocystis violacea 2311 a growth yield of 11.3g dry weight per mol thiosulfate consumed was obtained under chemoautotrophic conditions; under mixotrophic conditions with acetate the yield increased to 69g dry weight per mol thiosulfate consumed. With Thiocystis violacea 2311 maximal specific respiration rates were obtained with thiosulfate as electron donor irrespective of the presence or absence of sulfur globules in the cells; organic substrates served as carbon sources only and did not support respiration. With Chromatium vinosum D utilization of thiosulfate was not constitutive; maximal respiration rates on thiosulfate were obtained only with thiosulfate grown cells containing sulfur globules. Respiration rates were further increased by malate, fumarate or propionate; these substrates also served as sole electron donors for respiration. Acetate and pyruvate were used as carbon sources only. The ecological significance of the chemotrophic metabolism is discussed.     

Mixotrophic and Autotrophic Growth of Thiobacillus acidophilus on Glucose and Thiosulfate

Mixotrophic growth of the facultatively autotrophic acidophile Thiobacillus acidophilus on mixtures of glucose and thiosulfate or tetrathionate was studied in substrate-limited chemostat cultures. Growth yields in mixotrophic cultures were higher than the sum of the heterotrophic and autotrophic growth yields. Pulse experiments with thiosulfate indicated that tetrathionate is an intermediate during thiosulfate oxidation by cell suspensions of T. acidophilus. From mixotrophic growth studies, the energetic value of thiosulfate and tetrathionate redox equivalents was estimated to be 50% of that of redox equivalents derived from glucose oxidation. Ribulose 1,5-bisphosphate carboxylase (RuBPCase) activities in cell extracts and rates of sulfur compound oxidation by cell suspensions increased with increasing thiosulfate/glucose ratios in the influent medium of the mixotrophic cultures. Significant RuBPCase and sulfur compound-oxidizing activities were detected in heterotrophically grown T. acidophilus. Polyhedral inclusion bodies (carboxysomes) could be observed at low frequencies in thin sections of cells grown in heterotrophic, glucose-limited chemostat cultures. Highest RuBPCase activities and carboxysome abundancy were observed in cells from autotrophic, CO₂-limited chemostat cultures. The maximum growth rate at which thiosulfate was still completely oxidized was increased when glucose was utilized simultaneously. This, together with the fact that even during heterotrophic growth the organism exhibited significant activities of enzymes involved in autotrophic metabolism, indicates that T. acidophilus is well adapted to a mixotrophic lifestyle. In this respect, T. acidophilus may have a competitive advantage over autotrophic acidophiles with respect to the sulfur compound oxidation in environments in which organic compounds are present.     

Mixotrophic and heterotrophic growth of Beggiatoa alba in continuous culture

Beggiatoa alba strain B18LD was grown in continuous culture under heterotrophic conditions on acetate or acetate and asparagine and under mixotrophic conditions on acetate plus either 1 mM sodium sulfide or 1 mM sodium thiosulfate. Considerable differences were observed between the yields and the cell compositions of heterotrophic and mixotrophic cultures at all dilution rates tested. The dry weight yield per gram acetate utilized was approximately three times higher in the acetate-sulfide mixotrophic culture than in the acetate heterotrophic culture, whereas the poly--hydroxybutyric acid and carbohydrate contents were much higher in the heterotrophic cultures. The high yields (0.52–0.75, corrected for the weight of the sulfur) obtained with the mixotrophic cultures imply that the acetate was utilized mainly for biosynthesis. Thus, the oxidation of sulfide supplied energy. The addition of catalase to the chemostat cultures increased yields slightly, but it was insufficient to explain the differences between the heterotrophic and the mixotrophic cultures.     

Nitrogen fixation activity of a filamentous,nonheterocystous cyanobacterium in the presence and absence of exogenous,organic substrates

Gallionella ferruginea is able to utilize Fe(II) and the reduced sulfur compounds sulfide and thiosulfate as electron donor and energy source. Tetrathionate and elemental sulfur, on the other hand, are not metabolized. In sulfide-O₂ microgradient cultures G. ferruginea grows at the interface between the oxidizing and the reducing zones. Optimal growth depends on low oxygen and sulfide concentrations. Establishing within the gradient protects the bacterium from too high sulfide concentrations. G. ferruginea excretes extracellular polymeric substances (EPS). While in FeS-gradient cultures 2×10⁶ cells/ml were obtained the bacterial mass could be increased to 1–3×10⁸ cells/ml in shaken batch cultures using thiosulfate as substrate. A further increase of bacterial mass by adding an organic carbon source was not possible confirming that G. ferruginea is an obligate autotrophic organism. When growing on sulfide or thiosulfate the otherwise characteristic twisted stalk consisting of ferric hydroxide is lacking. It is thus shown to be a metabolic end product of Fe(II) oxidation rather than metabolically active cellular material.     

Magnetospirillum sulfuroxidans sp. nov., capable of sulfur-dependent lithoautotrophy and a taxonomic reevaluation of the order Rhodospirillales

A spiral-shaped, highly motile bacterium was isolated from freshwater sulfidic sediment. Strain J10^T is a facultative autotroph utilizing sulfide, thiosulfate, and sulfur as the electron donors in microoxic conditions. Despite high 16S rRNA gene sequence sequence identity to Magnetospirillum gryphiswaldense MSR-1 ^T (99.6 %), digital DNA-DNA hybridisation homology and average nucleotide identity between the two strains was of the different species level (25 % and 83 %, respectively). Strain J10^T is not magnetotactic. The DNA G + C content of strain J10^T is 61.9 %. The predominant phospholipid ester-linked fatty acids are C18:1ω7, C16:1ω7, and C16:0. Strain J10^T (=DSM 23205 ^T = VKM B-3486 ^T) is the first strain of the genus Magnetospirillum showing lithoautotrophic growth and is proposed here as a novel species, Magnetospirillum sulfuroxidans sp. nov. In addition, we propose to establish a framework for distinguishing genera and families within the order Rhodospirillales based on phylogenomic analysis using the threshold values for average amino acid identity at ̴ 72 % for genera and ̴ 60 % for families. According to this, we propose to divide the existing genus Magnetospirillum into three genera: Magnetospirillum, Paramagnetospirillum, and Phaeospirillum, constituting a separate family Magnetospirillaceae fam. nov. in the order Rhodospirillales. Furthermore, phylogenomic data suggest that this order should accomodate six more new family level groups including Magnetospiraceae fam. nov., Magnetovibrionaceae fam. nov., Dongiaceae fam. nov., Niveispirillaceae fam. nov., Fodinicurvataceae fam. nov., and Oceanibaculaceae fam. nov.     

Inactivation of the Flagellin Gene flaA in Magnetospirillum gryphiswaldense Results in Nonmagnetotactic Mutants Lacking Flagellar Filaments

Magnetotactic bacteria synthesize magnetosomes, which cause them to orient and migrate along magnetic field lines. The analysis of magnetotaxis and magnetosome biomineralization at the molecular level has been hindered by the unavailability of genetic methods, namely the lack of a means to introduce directed gene-specific mutations. Here we report a method for knockout mutagenesis by homologous recombination in Magnetospirillum gryphiswaldense. Multiple flagellin genes, which are unlinked in the genome, were identified in M. gryphiswaldense. The targeted disruption of the flagellin gene flaA was shown to eliminate flagella formation, motility, and magnetotaxis. The techniques described in this paper will make it possible to take full advantage of the forthcoming genome sequences of M. gryphiswaldense and other magnetotactic bacteria.     

Use of reduced sulfur compounds by Beggiatoa sp.

A strain of Beggiatoa cf. leptomitiformis (OH-75-B, clone 2a) was isolated which is unique among reported strains in its ability to deposit internal sulfur granules from thiosulfate. It also deposited these characteristic granules (as all BEggiatoa species do) from sulfide. In cultures where growth was limited by exhaustion of organic substrates, these granules generally comprised about 20% of the total cell weight. With medium containing acetate and thiosulfate, no measurable utilization of thiosulfate or deposition of elemental sulfur (S0) took place until after the exponential growth phase. Neither sulfide nor thiosulfate added an increment to heterotrophic growth yield except for the weight of the deposited S0. The deposition of S0 from thiosulfate was probably a disproportionation in which S0 and sulfate were produced in a 1:1 ratio. Some of the S0 was further oxidized to sulfate. No autotrophic or mixotrophic growth was demonstrated for this strain. When inoculated in small, well-dispersed quantities into yeast extract medium, this strain grew only after long lags. Addition of the enzyme catalase eliminated initial lags and increased growth rates slightly. In contrast, catalase had no influence on growth rate when added to mineral medium containing acetate. In yeast extract medium, the inhibition of growth rate was presumably because of peroxides. Addition of thiosulfate was almost as effective as catalase in eliminating this inhibition. The S0 granules which, in this case, were deposited during the exponential growth phase, appeared to be partly responsible for this relief. This strain of Beggiatoa sp. remained active for at least 5 days under strictly anaerobic conditions, and under those conditions, it increased its dry weight by about 2.5-fold. Anaerobic "growth" and maintenance required the presence of an energy source, such as acetate. When cells containing much internal S0 were transferred to an organic anaerobic medium, a substantial portion of the internal S0 was eventually converted to sulfide.     

Effects of light and temperature on the growth of Takayama helix (Dinophyceae): mixotrophy as a survival strategy against photoinhibition

Takayama helix is a mixotrophic dinoflagellate that can feed on diverse algal prey. We explored the effects of light intensity and water temperature, two important physical factors, on its autotrophic and mixotrophic growth rates when fed on Alexandrium minutum CCMP1888. Both the autotrophic and mixotrophic growth rates and ingestion rates of T. helix on A. minutum were significantly affected by photon flux density. Positive growth rates of T. helix at 6–58 μmol photons · m^?2 · s^?1 were observed in both the autotrophic (maximum rate = 0.2 · d^?1) and mixotrophic modes (0.4 · d^?1). Of course, it did not grow both autotrophically and mixotrophically in complete darkness. At ≥247 μmol photons · m^?2 · s^?1, the autotrophic growth rates were negative (i.e., photoinhibition), but mixotrophy turned these negative rates to positive. Both autotrophic and mixotrophic growth and ingestion rates were significantly affected by water temperature. Under both autotrophic and mixotrophic conditions, it grew at 15–28°C, but not at ≤10 or 30°C. Therefore, both light intensity and temperature are critical factors affecting the survival and growth of T. helix.     

Measuring magnetosomal pH of the magnetotactic bacterium Magnetospirillum magneticum AMB-1 using pH-sensitive fluorescent proteins

Magnetotactic bacteria synthesize uniform-sized and regularly shaped magnetic nanoparticles in their organelles termed magnetosomes. Homeostasis of the magnetosome lumen must be maintained for its role accomplishment. Here, we developed a method to estimate the pH of a single living cell of the magnetotactic bacterium Magnetospirillum magneticum AMB-1 using a pH-sensitive fluorescent protein E²GFP. Using the pH measurement, we estimated that the cytoplasmic pH was approximately 7.6 and periplasmic pH was approximately 7.2. Moreover, we estimated pH in the magnetosome lumen and cytoplasmic surface using fusion proteins of E²GFP and magnetosome-associated proteins. The pH in the magnetosome lumen increased during the exponential growth phase when magnetotactic bacteria actively synthesize magnetite crystals, whereas pH at the magnetosome surface was not affected by the growth stage. This live-cell pH measurement method will help for understanding magnetosome pH homeostasis to reveal molecular mechanisms of magnetite biomineralization in the bacterial organelle.     

Reactivity versus flexibility in thiobacilli

The results of ecophysiological studies on obligately and facultatively chemolithotrophic thiobacilli performed over the past years clearly show that the two types of organisms occupy different ecological niches. Chemostat experiments with cultures of the obligate chemolithotroph Thiobacillus neapolitanus and the facultative chemolithotroph Thiobacillus A2 have been carried out to explain the competitiveness of T. neapolitanus under conditions of strongly fluctuating substrate supply. Thiobacillus neapolitanus appeared to be very resistant to starvation periods whereafter it could oxidize sulfide (or thiosulfate) almost instantaneously at the original rate. Under alternate supply of 4 h sulfide and 4 h sulfate (or acetate which does not support growth of the organism either) to a chemostat culture of T. neapolitanus (D=0.05 h^–1) the sulfide concentration in the growth vessel never reached levels higher than 4m. This strategy is aimed at maximal reactivity. In contrast to T. neapolitanus the facultative chemolithotroph T.A2 appeared to be very flexible with respect to its energy generation. Under alternate supply of 4 h sulfide and 4 h acetate (D=0.05 h^–1) T.A2 was able to grow continuously since it directed its metabolism to either heterotrophy or autotrophy by rapid induction-repression mechanisms. This flexible strategy seems to be incompatible with a reactive strategy within one organism, since the oxidation capacity for sulfide decreased during the acetate period resulting in accumulation of sulfide during the sulfide period. It is concluded that T.A2 needs a continuous supply of an inorganic and an organic substrate to thrive whereas T. neapolitanus needs only a continuous supply of a reduced inorganic sulfur source but also will persist in environments with interrupted addition of sulfide provided that the starvation period does not last too long.     

Growth kinetics of haloalkaliphilic,sulfur-oxidizing bacterium<Emphasis Type="Italic"> Thioalkalivibrio versutus</Emphasis> strain ALJ 15 in continuous culture

The chemolithoautotrophic, sulfur-oxidizing bacterium Thioalkalivibrio versutus strain ALJ 15, isolated from a soda lake in Kenya, was grown in a continuous culture, with thiosulfate or polysulfide as growth-limiting energy source and oxygen as electron acceptor, at pH 10 and at pH 0.6, 2 M and 4 M total sodium. The end product of the sulfur-compound oxidation was sulfate. Elemental sulfur and a cell-bound, polysulfide-like compound appeared as intermediates during substrate oxidation. In the thiosulfate-limited culture, the biomass yields and maximum specific growth rates decreased two and three times, respectively, with increasing sodium concentration. The apparent affinity constant measured for thiosulfate and polysulfide was in the micromolar range (K_s=6±3 M). The maintenance requirement (m_s=8±5 mmol S₂O₃²/g dry weight h^–1) was in the range of values found for other autotrophic sulfur-oxidizing bacteria. The organism had a comparable maximum specific rate of oxygen uptake with thiosulfate, polysulfide, and sulfide, while elemental sulfur was oxidized at a lower rate. Glycine betaine was the main organic compatible solute. The respiration rates with different species of polysulfides (S_n^2–) were tested. All polysulfide species were completely oxidized at high rates to sulfate. Overall data demonstrated efficient growth and sulfur compounds oxidation of haloalkaliphilic chemolithoautotrophic bacteria from soda lakes.Communicated by W.D. Grant     

Energetics of mixotrophic and autotrophic C1-metabolism by Thiobacillus acidophilus

Although the facultatively autotrophic acidophile Thiobacillus acidophilus is unable to grow on formate and formaldehyde in batch cultures, cells from glucose-limited chemostat cultures exhibited substrate-dependent oxygen uptake with these C₁-compounds. Oxidation of formate and formaldehyde was uncoupler-sensitive, suggesting that active transport was involved in the metabolism of these compounds. Formate- and formaldehyde-dependent oxygen uptake was strongly inhibited at substrate concentrations above 150 and 400 M, respectively. However, autotrophic formate-limited chemostat cultures were obtained by carefully increasing the formate to glucose ratio in the reservoir medium of mixotrophic chemostat cultures. The molar growth yield on formate (Y=2.5 g ·mol^-1 at a dilution rate of 0.05 h^-1) and RuBPCase activities in cell-free extracts suggested that T. acidophilus employs the Calvin cycle for carbon assimilation during growth on formate. T. acidophilus was unable to utilize the C₁-compounds methanol and methylamine. Formate-dependent oxygen uptake was expressed constitutively under a variety of growth conditions. Cell-free extracts contained both dye-linked and NAD-dependent formate dehydrogenase activities. NAD-dependent oxidation of formaldehyde required reduced glutathione. In addition, cell-free extracts contained a dye-linked formaldehyde dehydrogenase activity. Mixotrophic growth yields were higher than the sum of the heterotrophic and autotrophic yields. A quantitative analysis of the mixotrophic growth studies revealed that formaldehyde was a more effective energy source than formate.     

Investigation of mixotrophic,heterotrophic, and autotrophic growth of Chlorella vulgaris under agricultural waste medium

Growth of Chlorella vulgaris and its lipid production were investigated under autotrophic, heterotrophic, and mixotrophic conditions. Cheap agricultural waste molasses and corn steep liquor from industries were used as carbon and nitrogen sources, respectively. Chlorella vulgaris grew remarkably under this agricultural waste medium, which resulted in a reduction in the final cost of the biodiesel production. Maximum dry weight of 2.62 g L^?1 was obtained in mixotrophic growth with the highest lipid concentration of 0.86 g L^?1. These biomass and lipid concentrations were, respectively, 140% and 170% higher than autotrophic growth and 300% and 1200% higher than heterotrophic growth. In mixotrophic growth, independent or simultaneous occurrence of autotrophic and heterotrophic metabolisms was investigated. The growth of the microalgae was observed to take place first heterotrophically to a minimum substrate concentration with a little fraction in growth under autotrophic metabolism, and then the cells grew more autotrophically. It was found that mixotrophic growth was not a simple combination of heterotrophic and autotrophic growth.     

Genetic improvement of <Emphasis Type="Italic">Magnetospirillum gryphiswaldense</Emphasis> for enhanced biological removal of phosphate

Objectives

To improve its phosphate accumulating abilities for phosphate recycling from wastewater, a magnetotactic bacterium, Magnetospirillum gryphiswaldense, was genetically modified to over-express polyphosphate kinase.

Results

Polyphosphate kinase was over-expressed in the bacterium. The recombinant strain accumulated ninefold more polyphosphate from synthetic wastewater compared to original wild type. The magnetic property of the recombinant M. gryphiswaldense strain was retained.

Conclusions

The recombinant M. gryphiswaldense can be used for phosphate removal and recovery in bioremediation.

     

Effect of organic carbon,C:N ratio and light on the growth and lipid productivity of microalgae/cyanobacteria coculture

Current culture methods based on monocultures under phototrophic regimes are prone to contamination, predation, and collapse. Native cultures of multiple species are adapted to the local conditions and are more robust against contamination and predation. Growth, lipid and biomass productivity of a Louisiana native coculture of microalgae (Chlorella vulgaris) and cyanobacteria (Leptolyngbya sp.) in heterotrophic and mixotrophic regimes were investigated. Dextrose and sodium acetate at C:N ratios of 15:1 and 30:1 under heterotrophic (dark) and mixotrophic (400 μmol m^?2 s^?1) regimes were compared with autotrophic controls. The carbon source and C:N ratio impacted growth and biomass productivity. Mixotrophic cultures with sodium acetate (C:N 15:1) resulted in the highest mean biomass productivity (156 g m^?3 d^?1) and neutral lipid productivity (24.07 g m^?3 d^?1). The maximum net specific growth rate (U) was higher (0.97 d^?1) in mixotrophic cultures with dextrose (C:N 15:1) but could not be sustained resulting in lower total biomass than in mixotrophic cultures with acetate (C:N 15:1), with a U of 0.67 d^?1. The ability of the Louisiana coculture to use organic carbon for biomass and lipid production makes it a viable feedstock for biofuels and bioproducts.     

Competition between the facultatively chemolithotrophic Thiobacillus A2, an obligately chemolithotrophic Thiobacillus and a heterotrophic spirillum for inorganic and organic substrates

Competition in a chemostat between the versatile Thiobacillus A2 and the specialized T. neapolitanus for thiosulfate as the sole growth-limiting substrate, led to dominance of the specialized over the versatile organism, at dilution rates 0.025 h^-1. Increasing concentrations of acetate or glycollate in the thiosulfate medium caused increased relative numbers of T. A2 in steady states at D=0.07 h^-1. Eventually, with 10–12 mmol of organic substrate per litre, complete dominance of T. A2 over T. neapolitanus occurred.Mixed cultures of T. A2 and a specialized spirillumshaped heterotroph, competing for acetate as sole growth-limiting substrate resulted in complete dominance of the heterotroph at dilution rates of 0.07 and 0.15 h^-1. In this case increasing concentrations of thiosulfate in the acetate medium, up to 10 mM, eventually led to the elimination of the heterotroph.These results have been interpreted as evidence that T. A2 was growing mixotrophically. As the concentration of the second substrate was raised, the number of T. A2 cells increased and as a result T. A2 consumed an increasing portion of the common substrate.In mixed chemostat cultures containing all three organisms, T. A2 could maintain itself with all tested ratios of acetate and thiosulfate in the inflowing medium. The heterotroph was excluded from the culture below a relatively low acetate to thiosulfate ratio, whilst above a relatively high acetate to thiosulfate ratio T. neapolitanus was completely eliminated.These results were discussed in relation to the ecological niche of Thiobacillus A2-type organisms.     

Growth yields of Desulfotomaculum orientis with hydrogen in chemostat culture

Desulfotomaculum orientis (strain Singapore 1) was grown autotrophically with H₂+CO₂ and sulfate, thiosulfate or sulfite as electron acceptor in sulfide- and pH-controlled continuous culture. Under sulfate-limiting conditions real growth yields of up to 9.7 g cell dry mass per mol sulfate were obtained. Electron acceptor limitation resulted in the excretion of up to 14.5 mmol acetate per liter, formed by reduction of CO₂ with H₂. Acetate production was not coupled to an increase of growth yields: under hydrogen-limiting conditions only 1.6 mmol acetate per liter was produced, and even higher growth yields of up to 12,4 g cell dry mass per mol sulfate were obtained. With thiosulfate or sulfite as electron acceptor growth yields increased up to 17.9 g cell dry mass per mol electron acceptor. Growth yields were not simply correlated with the growth rate, and did not allow the determination of maintenance coefficients and the extrapolation to maximal yields at infinite growth rate (Y ^max). The maximal growth rates (_max) with sulfate and thiosulfate were 0.090 and 0.109 h^-1, respectively, if cells were grown continuously in sulfidostat culture under nonlimiting conditions.The net energy yield of sulfate reduction and the energy requirement for the activation of sulfate by Desulfotomaculum orientis are discussed.